Xia et al.

Horticulture Research (2020)7:7

https://doi.org/10.1038/s41438-019-0225-4
Horticulture Research
www.nature.com/hortres

REVIEW ARTICLE Open Access

Tea plant genomics: achievements, challenges and

perspectives
En-Hua Xia1, Wei Tong1, Qiong Wu1, Shu Wei1, Jian Zhao1, Zheng-Zhu Zhang1, Chao-Ling Wei1 and Xiao-Chun Wan1

Abstract
Tea is among the world’s most widely consumed non-alcoholic beverages and possesses enormous economic, health,
and cultural values. It is produced from the cured leaves of tea plants, which are important evergreen crops globally
cultivated in over 50 countries. Along with recent innovations and advances in biotechnologies, great progress in tea
plant genomics and genetics has been achieved, which has facilitated our understanding of the molecular
mechanisms of tea quality and the evolution of the tea plant genome. In this review, we briefly summarize the
achievements of the past two decades, which primarily include diverse genome and transcriptome sequencing
projects, gene discovery and regulation studies, investigation of the epigenetics and noncoding RNAs, origin and
domestication, phylogenetics and germplasm utilization of tea plant as well as newly developed tools/platforms. We
also present perspectives and possible challenges for future functional genomic studies that will contribute to the
acceleration of breeding programs in tea plants.
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Introduction transcriptome of tea plants using diverse NGS technolo-

The tea plant (Camellia sinensis) is an economic crop
with significant importance1. Its leaves are generally used
to produce tea—the world’s most popular non-alcoholic
beverage. Tea comprises abundant characteristic com-
pounds, such as tea polyphenols, theanine, and caffeine,
which not only determine the quality of tea but also
confer it with tremendous health benefits2. However,
compared to other crops, such as rice3, research on the
functional genomics of tea plants is lagging behind, which
has hampered the utilization of genetic resources for
modern molecular breeding, which is urgently needed.
The rapid development and innovation of biotechnolo-
gies, particularly next-generation sequencing (NGS), has
resulted in significant progress in understanding the
genomics and genetics of tea plants in the past two dec-
ades, which can be briefly summarized according to five
aspects: (1) the deciphering of the genome and
Correspondence: Chao-Ling Wei (weichl@ahau.edu.cn) or
Xiao-Chun Wan (xcwan@ahau.edu.cn)
1 tury. Morinaga and colleagues observed that 15 chromo-
State Key Laboratory of Tea Plant Biology and Utilization, Anhui Agricultural
University, Hefei 230036, China
These authors contributed equally: En-Hua Xia, Wei Tong
gies; (2) the identification and functional analysis of tea
quality-related genes and their regulatory networks; (3)
the investigation of the epigenetics and noncoding RNAs
(ncRNAs) involved in stress responses and other biolo-
gical processes; (4) the exploration of the diversification
and domestication of tea plants; and (5) the development
of innovative tools and techniques for the characterization
of novel genes and alleles. In this review, we briefly
summarize the main achievements of the last two decades
and present potential challenges and perspectives for
future functional genomic studies that would contribute
to the acceleration of breeding programs in tea plants.
Whole-genome sequencing of tea plants
Cytogenetics of tea plants
Tea plants are an economically important crop. Exten-
sive investigations of tea plant karyotypes have facilitated
our fundamental understanding of their chromosomal
biology4–7. The first report of the chromosome karyotype
analysis of tea plants can be traced back to the last cen-
somes (n = 15) were present in the gametes of C. sinensis

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Xia et al. Horticulture Research (2020)7:7
tea plants and that 30 chromosomes (2n = 30) occurred in
the zygotes of C. japonica, a closely related species of tea
plant8,9. These findings suggested that the diploid tea
plant has 30 chromosomes and that the chromosome
number might be conserved in genus Camellia. New
innovations in biotechnologies gradually confirmed these
conclusions among globally collected tea clones5,7,10–12.
In particular, the investigation of the chromosome num-
bers and ploidy levels of 139 tea individuals representing
almost all sections of genus Camellia revealed the diploid
nature of the majority (90.65%) of Camellia species, with a
chromosome number of 2n = 306. Only a few tea plant
species are polyploid, which exhibit 45–120 chromo-
somes. These species predominantly come from section
Camellia (C. chekiangoleosa, C. mairei, and C. reticulata),
section Oleifera (C. oleifera and C. sasanqua), and section
Archecamellia (C. granthamiana).
Genome size refers to the DNA content within the
haploid genome of an organism. The estimation of gen-
ome size is not only of great significance for cytogenetic
studies but also provides indispensable basic data for
genome sequencing, comparative genomics, and evolu-
tionary analyses. The genome size of tea plants was
initially estimated to be 3.8–4.0 Gb13,14, which was further
demonstrated by the investigation of 36 Indian tea
accessions, where all the investigated accessions exhibited
diploid genomes with sizes ranging from 3.5 to 3.8 Gb15.
The recent genome sequencing of two representative elite
tea plant cultivars, C. sinensis var. sinensis cv. shuchazao
and C. sinensis var. assamica cv. yunkang#10, showed an
~3.0 Gb genome size, which is similar to that of maize but
much larger than those of coffee and cocoa1,2. While
genome size is likely to be conserved in cultivated tea
plants, intraspecific and interspecific variations are also
observed in genus Camellia, possibly due to the frequent
hybridization and polyploidization events that occur after
speciation6.
Genetic linkage maps of tea plants
The construction of genetic linkage maps is the basis of
molecular biology and is essential for a wide range of
genetics and genomic studies, such as quantitative trait
mapping, molecular marker-assisted breeding and com-
parative genomic studies16. Unlike other crops, tea plants
are self-incompatible species with relatively low cross-
fertility rates, resulting in a lack of a high generation
segregation populations and sufficient offspring for
genetic map construction17. More than 10 genetic linkage
maps have been generated for tea plants, most of which
are produced based on an F1 population consisting of
<150 offspring18–22 (Table 1). The dominant molecular
markers used for genetic map construction include ran-
dom amplification of polymorphic DNA (RAPD) mar-
kers18,20,23–26, amplified fragment length polymorphism
Page 2 of 19
(AFLPs)18–20, inter-simple sequence repeats (ISSRs)24,
simple sequence repeats (SSRs)20,25–28, and single
nucleotide polymorphisms (SNPs)21,29. These reported
genetic maps vary in length and density, with the total
map length ranging from 1180.9 to 3314.3 cM and the
average distance between adjacent markers from 0.4 to
20.1 cM. Such differences are principally affected by the
total number of individuals and types of molecular mar-
kers applied for map construction. The highest-density
genetic map of tea plants obtained to date was con-
structed using a total of 4217 polymorphic SNP markers
developed from 327 individuals of the F1 segregating
population (LJ43 × BHZ) via 2b-RAD sequencing29. The
resultant genetic maps consisted of 15 linkage groups
ranging from 87.46 to 146.93 cM. The total map length
was 1679 cM, with an average interval of 0.4 cM between
4217 adjacent markers. LG01 and LG03 were the two
largest linkage groups, with genetic distances of
146.93 cM (374 markers) and 146.61 cM (374 markers),
respectively. Despite the numerous genetic maps cur-
rently available, the density and quality of these maps
need to be further improved by investigating larger
populations and applying more advanced biotechnologies.
The resultant data will be important for future investi-
gations of tea plant biology that will benefit the whole tea
industry.
Current progress in tea plant genome sequencing
With the revolution of sequencing technology, an
increasing number of labs around the world have suc-
cessfully released more than 236 plant genomes, and the
tea plant genome is probably among the most complex of
these30 (Fig. 1). This is chiefly because tea plants have an
extremely large, highly heterozygous, repeat-rich nuclear
genome, presenting major challenges in genome assem-
bly. The tea community has put forth its best effort to
assemble and release the genomic sequences of two pri-
mary tea plant varieties: C. sinensis var. sinensis (CSS;
Chinese type tea) and C. sinensis var. assamica (CSA;
Assam type tea), using NGS technologies1,2. Both of the
assemblies are composed of ~3 Gb of genomic sequences
with an average scaffold N50 size of 920 Kb, which is
much larger than those from other plants with complex
genomes, such as the orchid (scaffold N50 = 359 Kb)31
and moso bamboo (scaffold N50 = 329 Kb)32. Never-
theless, compared to other model species, such as rice33,
the scaffold N50 sizes of the current assemblies are still
somewhat disappointing, due largely to the limitation
imposed by the short read length generated from NGS
technology (Fig. 1). Unassembled regions account for ~5%
of the current assemblies. In the future, additional efforts
will be needed to further improve the quality and com-
pleteness of the genome assemblies of tea plants, parti-
cularly with the upcoming advances in sequencing
Xia et al. Horticulture Research (2020)7:7 Page 3 of 19

Table 1 Summary of the representative linkage maps of tea plants.

Mapping population Sizea Marker type No. of Total Marker No. of Reference
markers length (cM) density (cM) linkage groups

Sayamakaori × Kana-CK17 54 RAPD 140 1640 11.7 17 Ota et al.23

SFS150 × TN14/3 90 RAPD 126 1350 11.7 15 Hackett et al.18
AFLP
Keemun #4 × 69 AFLP 208 2458 11.9 17 Huang et al.19
Chaoandawuye
Fudingdabaicha × 94 ISSR 62 1181 20.1 7 Huang et al.24
Zhenong #12 RAPD
SFS150 × S15/10 42 SSR 100 1412 14.1 30 Kamunya et al.20
RAPD
AFLP
Sayamakaori × Kana-CK17 54 SSR 279 1218 4.35 15 Taniguchi et al.25
STS
CAPS
RAPD
Yingshuang × 183 SSR 406 1144 2.9 15 Ma et al.27
Beiyuedanzhu
Yingshuang × 148 SSR 6448 3965 1.0 15 Ma et al.21
Beiyuedanzhu SNP
Long #43 × Baihaozao 170 SSR 483 1226 2.5 15 Tan et al.28
Fushun × Kemsull 79 SSR 678 1442 4.7 15 Chang et al.26
RAPD
AFLP
Long #43 × Baihaozao 327 SNP 4217 1679 0.4 15 Xu et al.29
a
Population size

technologies (e.g., 3rd-generation sequencing) and new
algorithm-derived assemblers.
There are 33,932 and 36,951 protein-coding genes in
the CSS and CSA tea plant genomes, respectively.
Transposable elements (TEs) account for 64% and 80% of
the CSS and CSA genomes, respectively. LTR retro-
transposons are among the most dominant TEs, inde-
pendently representing 58% and 67% of the CSS and CSA
genomes, respectively. The rapid propagation of TEs,
particularly the proliferation and persistence of a single
Ty3/gypsy retrotransposon family (TL001) for over 50
million years, has been proven to have led to a con-
siderable increase in genome size in tea plants1. CSA and
CSS diverged from their common ancestor ~0.38–1.54
million years ago (mya). However, this divergence time
might have been underestimated because of the draft
nature of the two current genome assemblies, which
generated only a small proportion of collinear genes for
dating. Similar to other plants, tea plants underwent two
whole-genome duplication (WGD) events, including
recent and ancient events occurring 30–40 and 90–100
mya, respectively2.
Tea plants contain rich and diverse secondary meta-
bolites that not only confer tea quality characteristics,
such as color, aroma and taste, but also play critical roles
in the responses to biotic and abiotic stresses34. Several
genes encoding enzymes associated with the biosynthesis
of secondary metabolites were significantly amplified in
the tea plant genome. In particular, the striking expansion
of serine carboxypeptidase-like (SCPL) acyltransferase-
encoding genes might directly contribute to the high
accumulation of galloylated catechins that determine tea
palatability2. Compared to kiwifruit, tomato, potato,
cacao, and Arabidopsis thaliana, tea plants show expan-
sion of disease resistance genes, including nucleotide-
binding sites with leucine-rich repeats (NBS-LRRs) and
pattern-recognition receptors (RLK-LRRs)1. This con-
tributes to the enhancement of the immune system of tea
plants and, thus, their adaptation to diverse global
environments. WGD events and subsequent tandem
Xia et al. Horticulture Research (2020)7:7 Page 4 of 19

4
Capsicum baccatum
Helianthus annuus
Capsicum annuum Roche 454

Capsicum chinense Illumina

Camellia sinensis var. assamica Chinese Oxford Nanopore
3 type tea
PacBio
Camellia sinensis var. sinensis
Sanger
Estimated genome size (Gb)

Assam type tea Zea mays (CML247) Zea mays B73)

Nicotiana attenuata

2 Gossypium hirsutum

Elaeis guineensis Coffea canephora Chenopodium quinoa

Actinidia chinensis
Brassica oleracea
1 Sorghum bicolor
Theobroma cacao
Populus trichocarpa Vitis vinifera CS
Rosa chinensis
Oryza meridionalis Vitis vinifera

Oryza sativa ssp. indica

0 Prunus persica Arabidopsis thaliana

10k 100k 1M 10M

Contig N50 of genome assembly

Fig. 1 Current genome sequencing progress in tea and other plants. The x-axis represents the contig N50 of the genome assembly, while the y-
axis shows the estimated genome size of each plant. The sequencing platforms are indicated in red (Roche 454), brown (Illumina), green (Oxford
nanopore), blue (PacBio SMRT), and pink (Sanger). Tea plants are highlighted with a rectangular box.

duplications are the two major evolutionary forces that methyltransferases (NMTs)35. The NMT genes have
drive such gene expansions. undergone recent rapid and independent evolution in tea
Tea-processing suitability refers to the characteristics of plants relative to cocoa and coffee1.
tea varieties that make them suitable for the manufacture
of certain types of tea to achieve the best quality. It is Organelle genome sequencing
accepted that different varieties of tea plants differ in Plants generally harbor two independent organelle
terms of flavors and tastes. The sequencing of the tea genomes (chloroplast and mitochondria), which provide
plant genome together with the transcriptomes and invaluable resources for a range of functional, evolu-
metabolomes of 25 Camellia species representing almost tionary and comparative genomic studies36. The culti-
all the sections from genus Camellia showed that, despite vated tea plant harbors a circular chloroplast (cp) genome
the conservation of three metabolic pathways (catechins, of 157,096 bp in length with an overall GC content of
theanine, and caffeine) among Camellia species, most of 37.3%37. It exhibits the typical plant cp genome structure,
the flavonoid and caffeine, but not the theanine-related including a pair of inverted repeat regions (IRs, 26,080 bp)
genes, exhibited higher expression levels in species from separated by a large single-copy region (LSC, 86,653 bp)
section Thea than in non-Thea species1. This elucidates and a small single-copy region (SSC, 18,283 bp). The
why the leaves of tea plants from non-Thea sections, such whole-genome sequence encodes a total of 133 cp genes,
as well-known ornamental camellias (e.g., C. japonica) 86 of which are protein-coding genes, while 8 are rRNA
and the traditional oil tea plant (C. oleifera), accumulate genes, and 39 are tRNA genes. The cp genomes have been
such low contents of catechins and caffeine but not assembled for diverse well-known tea cultivars, such as
theanine, making them unsuitable for tea production. “Longjing #43”37, “Yunkang #10”38, and “Shuchazao”2. In
Further investigation of the roles of natural selection and addition to cultivated tea plants, an increasing number of
artificial selection in shaping the genes involved in the cp genomes from species of genus Camellia have also
three metabolic pathways will contribute to a deep been released, including those of C. japonica
understanding of the processing suitability and quality of (NC_036830), C. mairei (NC_035688), C. szechuanensis
tea. In addition, caffeine (1,3,7-trimethylxanthine) is (NC_035651), C. elongata (NC_035652), C. azalea
among the most well-known purine alkaloids in plants, (NC_035574), C. pubicosta (NC_024662), C. petelotii
and its biosynthesis is mainly catalyzed by the N- (NC_024661), C. reticulata (NC_024663), C. grandibracteata
Xia et al. Horticulture Research (2020)7:7
(NC_024659), C. leptophylla (NC_024660), C. crapnelliana
(NC_024541), C. oleifera (NC_023084), C. danzaiensis
(NC_022460), C. impressinervis (NC_022461), C. yunna-
nensis (NC_022463), C. cuspidata (NC_022459), C.
pitardii (NC_022462), and C. taliensis (NC_022264)39–45.
The Camellia cp genomes possess quite similar genome
sizes, total numbers of genes, and sequence homology,
suggesting extreme genome conservation during
evolution.
In sharp contrast to the chloroplast genome, only a
partially assembled mitochondrial (mt) genome is avail-
able for cultivated tea plants38. The current release of the
tea plant mt genome consists of two circular scaffolds
with total lengths of 702,253 bp (45.63% GC) and
178,082 bp (45.81% GC). It encodes 71 mt genes,
including 44 protein-coding genes, 24 tRNA genes, and 3
rRNA genes. Compared to the cp genome, the tea plant
mt genome shows a repeat-rich nature, including a total
of 38,027 bp of long repeat sequences38.
Transcriptome sequencing and gene discovery in
tea plants
Transcriptome sequencing of tea plants
Transcriptome sequencing has revolutionized genetic
and functional genomic studies of organisms, particularly
for nonmodel species without available sequenced gen-
omes46. In the last decade, innovations overcoming
challenges in tea plant genome sequencing have greatly
accelerated the transcriptomic investigation of this eco-
nomically important crop. Shi and colleagues examined
the first transcriptome of tea plants (cultivar shuchazao)
using Illumina sequencing technology and obtained a
total of 127,094 unigenes, which were applied for the in-
depth exploration of candidate genes involved in the
biosynthesis of characteristic compounds of tea plants
that determine tea quality47. Subsequently, growing
transcriptome-sequencing projects were launched to fur-
ther investigate the gene expression dynamics of tea
plants under cold acclimation48–51, drought stress52,53,
and hormone responses54 and the mechanisms underlying
self-incompatibility17,55, nitrogen utilization56,57, trichome
formation58, and tea quality59–62. These results greatly
broadened our understanding of tea plant biology. With
the advent of single-molecule sequencing technology, a
recent study produced a more accurate full-length tran-
scriptome of tea plants63. In addition to the cultivated tea
plants, several transcriptomes from closely related species
from genus Camellia have also been reported, which have
provided indispensable resources for comparative tran-
scriptomic studies64–66 (Supplementary Table 1). The
sequencing of the transcriptome of oil tea plants (C.
oleifera) identified 3022 orthologous gene pairs between
cultivated tea plants and oil tea plants, among which 211
exhibited evidence of positive selection64. Compared to
Page 5 of 19
the cultivated tea plants, C. taliensis showed extraordinary
amplification of cold tolerance-related genes65. Most of
the genes associated with triacylglycerol biosynthesis in C.
reticulata and C. sinensis are multiple-copy genes, sug-
gesting the potential occurrence of WGD events in the
common ancestor of genus Camellia66. An increasing
number of transcriptome-sequencing projects have also
been carried out in other Camellia species, such as C.
sasanqua67, C. chekiangoleosa68, and C. nitidissima69.
These datasets expanded the gene pool of tea plants and
will be of particular importance for future tea plant
breeding, as well as investigations of functional genomics,
phylogenomics, and comparative transcriptomics. It
should be noted that the current reported transcriptomes
from both tea plants and other Camellia species vary in
the total number of assembled transcripts and N50 sizes,
primarily due to the different sequencing depths and
assemblers adopted. In the future, more efforts should be
made to evaluate the optimized-sequencing depth and
assembler to better construct the transcriptomes of tea
plants.
Functional gene sets and their regulatory networks
One fundamental mission of the molecular biology
research on tea plants is to understand the functions and
regulatory mechanisms of genes encoded by the genome.
During recent decades, rapid advances in biotechnologies
have facilitated the cloning and functional characteriza-
tion of an increasing number of genes in tea plants70–72.
Most of these genes have been identified from CSS and
CSA, in which the numbers of cloned genes are much
greater than those from other closely related tea plant
species. Their functions can generally be classified into
three major categories, including secondary metabolite
biosynthesis73–80, abiotic and biotic stress responses81–86,
and aroma formation70,71,81,87–91 (Table 2). Among these
genes, those associated with secondary metabolite bio-
synthesis and aroma formation are the most studied
because they directly determine the quality of tea. Sup-
plementary Table 2 summarizes the details of the cur-
rently available cloned genes in tea plants. We here
selectively highlight some representatives that would be
preferred candidates for the future genetic improvement
of tea plants.
Caffeine is the most abundant purine alkaloid in the
majority of tea plants99. It is a crucial component of tea
quality and is significantly related to the bitterness of tea.
Nevertheless, excessive intake of caffeine has been
reported to have some side effects for human health, such
as increasing the risk of cardiovascular disease, palpita-
tions, and insomnia100,101. The documentation of the
genes controlling caffeine biosynthesis is therefore
essential for the future of breeding new varieties with a
low caffeine content. Kato et al.73 cloned the gene coding
Xia et al. Horticulture Research (2020)7:7 Page 6 of 19

Table 2 List of representative functional genes in tea plants.

Gene symbol NCBI accession Length (bp) Function description Ref.

Polyphenol biosynthesis
77
CsANR1 GU992402 1044 Encoding anthocyanidin reductase
77
CsANR2 GU992400 1014 Encoding anthocyanidin reductase
CsLAR GU992401 1282 Involved in flavan-3-ol biosynthesis 77

CsF3'H KT180309 1706 Encoding flavonoid 3'-hydroxylase 92

CsC4H KY615675 1518 Encoding cinnamate 4-hydroxylase in synthesis of flavonoids and lignins 93

CsF3H KY615688 1107 Encoding flavanone-3-hydroxylase 94

78
CsMYB4a KY774676 735 Negatively regulates phenylpropanoid and shikimate pathway
79
CsMYB5 KY827396 930 Regulate anthocyanin and proanthocyanidin biosynthesis
76
CsMYB75 GGTM01017033 765 Regulate anthocyanin hyperaccumulation
95
CsGST MK431867 642 Encoding glutathione-S-transferase
Caffeine biosynthesis
73
CsTCS AB031280 1438 Caffeine synthase gene
Theanine biosynthesis
CsTSI TEA015198a 2544 Theanine synthetase gene 2

CsAAPs TEA031577a 2339 Encoding amino acid permease involved in theanine transportation 96

Aroma formation
Csβ-PD AB088027 1729 Encoding beta-primeverosidase in floral aroma formation 88

CsGT1 AB847092 1458 Converting volatile organic compounds into β-primeverosides for aroma biosynthesis 90

97
CsSAMT MG459470 1104 Encoding the salicylic acid carboxyl methyltransferase for methyl salicylate formation
87
CsLIS/NES KF006849 2050 Producing (E)-nerolidol and linalool in vitro
81
CsUGT85A53 NA 1455 Formation of (Z)‐3‐hexenyl glucoside, novel insect pest control
Abiotic and biotic stress
82
CsCOR1 EU563236 755 Involved in cold, salinity and dehydration tolerance
98
CsHSP17.2 KU244518 734 Mediates tea plant thermo tolerance
84
CsICE1 GQ229032 1964 Inducer of CBF expression
84
CsCBF1 EU563238 1210 C-repeat-binding factor
83
CsTLP DQ444296 681 Increases tolerance to fungal pathogens overexpressed in potato
86
CsGolS2 KP757767 1574 Response to Ectropic oblique attack
a
CSS locus ID (from TPIA: http://tpia.teaplant.org)

caffeine synthase (TCS) from the young leaves of tea
plants using the rapid amplification of complementary
DNA ends (RACE) technique73. This gene is 1438 bp in size
and encodes 369 amino acids. The expression of the TCS
gene in Escherichia coli enables high production of caffeine
after feeding on xanthine and S-adenosylmethionine as
substrates, confirming the caffeine synthase activity of TCS.
Theanine is another characteristic amino acid of tea plants
and is related to the fresh tastes of tea. The gene encoding
enzyme associated with theanine biosynthesis (CsTSI) was
identified through a combination of genomic and
transcriptomic analyses2. This gene shares high homology
with the glutamine synthetase (PtGS) gene of Pseudomonas
taetrolens, indicating its potential bacterial origin102. The
overexpression of CsTSI in A. thaliana significantly increa-
ses the accumulation of theanine after ethylamine feeding.
Tea polyphenols are the major secondary metabolites of tea
plants103 and are closely related to the astringent and bitter
taste of tea. Two enzymes, UDPglucose:galloyl-1-O-β-D-
glucosyltransferase (UGGT) and epicatechin:1-O-galloyl-β-
D-glucose O-galloyltransferase (ECGT), were purified and
shown to be involved in the biosynthesis of galloylated
Xia et al. Horticulture Research (2020)7:7
catechins74. Three UDP-glycosyltransferase (UGT) genes,
CsUGT84A22, CsUGT78A14, and CsUGT78A15, were
found to be involved in the biosynthesis of β-glucogallin and
glycosylated flavonols75. Anthocyanin is another type of
polyphenol that typically accumulates in purple tea varieties.
It has considerable health benefits and has been used to
develop anthocyanin-enriched beverages. The expression of
a gene encoding leucoanthocyanidin reductase (CsLAR) in
E. coli results in the production of 2R,3S-trans-flavan-ol
(+)-catechin after the feeding of leucocyanidin as a sub-
strate77. Similarly, the expression of two anthocyanidin
reductase (CsANR1 and CsANR2)-encoding genes in E. coli
enables the conversion of cyanidin to a mixture of
(+)-epicatechin and (−)-catechin77. More recently,
CsMYB75 and CsGSTF1 were found to be involved in
anthocyanin accumulation in purple tea76. CsMYB75 can
promote the expression of CsGSTF1 in transgenic tobacco
plants. CsGSTF1 enables the transfer of anthocyanin from
the endoplasmic reticulum (ER) to vacuoles76. In tea plants,
the regulation of the catechin metabolic pathway is complex,
and several transcription factors (e.g., MBW complexes) are
involved76,78–80,104.
Aroma is vital for tea quality and for attracting global
interest. Since the advent of new chemical analytical
techniques, such as mass spectrometry, considerable
efforts have been made to identify volatile constituents of
different types of teas and to assess volatile odor activities
and their contribution to tea aroma70,71,88,89,105–109. Sig-
nificant studies on tea aroma biology have revealed that
hundreds of volatile terpenoids, in addition to some other
volatiles, such as cis-3-Hexen-1-ol, are present in tea as
glycosides110–112, which can be released during the tea
manufacturing process113. A tea beta-primeverosidase
gene was isolated and expressed in E. coli. The prokar-
yotically expressed mature protein was found to be able to
hydrolyze beta-primeverosides, liberating a primeverose
unit and aglycons and playing roles in tea plant defense
and floral aroma formation during the tea manufacturing
process88. Two UGTs from C. sinensis, UGT85K11
(CsGT1) and UGT94P1 (CsGT2), were found to convert
volatiles into β-primeverosides via sequential glucosyla-
tion and xylosylation, respectively90. More recently, a
UGT gene (CsUGT85A53) that was functionally validated
was shown to enable the conversion of exogenous (Z)-3-
hexenol from damaged adjacent tea leaves to its glucoside
form, enhancing the ability of tea plants to defend against
pests81. Studies on tea volatile biosynthesis have also
revealed an interesting bifunctional linalool/nerolidol
synthase gene (CsLIS/NES) in tea plants87. This gene
transcribes two transcript isoforms, CsLIS/NES-1 and
CsLIS/NES-2, which possess distinct subcellular locali-
zations and molecular functions, with CsLIS/NES-1
localizing to chloroplasts and functioning as linalool
synthase, while CsLIS/NES-2 localizes to the cytosol and
Page 7 of 19
potentially acts as a nerolidol synthase. In addition, the
biosynthesis of the tea volatiles indole, linalool, and ner-
olidol was found to be controlled by the corresponding
genes in tea89,91,114. Collectively, the aforementioned
genes that were cloned and functionally validated in tea
plants have broadened our understanding of the genetic
basis of tea quality and will be particularly useful for the
future genetic improvement of tea plants.
DNA methylation of tea plants
DNA methylation is among the most essential and
ubiquitous epigenetic modifications in plants115. It plays
crucial roles in gene expression regulation, cell differ-
entiation, and transposon element (TE) silencing116. DNA
methylation are mostly investigated in model plants, such
as Arabidopsis and rice, due to their small genome sizes
and low genome complexity117,118. Tea plants are non-
model woody plants with complex nuclear genomes. The
sequencing of the tea plant genome has revealed an
extraordinary amplification of genome size driven pri-
marily by a burst of TEs1,2. DNA methylation was con-
firmed to be closely linked with TE burst in tea plants119.
Compared to asterids such as potato, tea plants exhibit
higher CHG methylation levels, similar to those found in
maize and Norway spruce, two plant species with a
comparable genome size and repeat content to tea
plants120,121. Remarkably, the DNA methylation levels of
TEs vary with their evolutionary distance in tea plants,
with the methylation level increasing in recent, active TEs
and decreasing in ancient TEs. It is widely accepted that
DNA methylation can spread from TE boundaries to close
genomic regions, resulting in increased methylation levels
of adjacent genes122. High TE contents in tea plants might
therefore contribute to the high level of DNA methyla-
tion. Several genes involved in the biosynthesis of sec-
ondary metabolites (catechins, theanine, and caffeine)
were found to be TE-related in tea plants. This suggests
that TE-mediated DNA methylation may have some
effects on the formation of tea quality.
Chilling damage caused by low temperature occurs
commonly in tea plants and has severely affected the
sustainable development of the global tea industry.
Approximately 49–51% of cytosine residues are methy-
lated during the cold acclimation of tea plants123. Com-
pared to preacclimation, the DNA methylation level
increases significantly during cold acclimation and is
maintained at a higher level after deacclimation. The gene
encoding DNA methyltransferase, CsDRM2, was cloned
from an elite cultivar of tea plant (Longjing #43) and
found to exhibit high expression levels under cold accli-
mation124. In the future, additional DNA methylation
projects need to be performed in relation to other aspects
of tea plants to elucidate the roles of these modifications
in plant development, environmental adaptation, stress
Xia et al. Horticulture Research (2020)7:7
responses, and even genome structure variation and
stabilization.
ncRNAs of tea plants
MicroRNAs (miRNAs), a class of endogenous small
ncRNAs, have been recognized as a critical genetic reg-
ulator primarily engaged in posttranscriptional regulation.
In tea plants, a series of studies have been undertaken to
examine the posttranscriptional regulation of miRNAs in
development125,126, metabolism126–128, and responses to
biotic/abiotic stresses129–133. Tea plant miRNAs can
negatively regulate catechins and terpenoid biosynthesis
by down-regulating target genes related to their bio-
synthesis at both the transcriptional and posttranscrip-
tional levels126,128. For instance, Cs-miR156 regulates
catechin accumulation in tea plants by suppressing the
expression of the target gene SPL in the presence of dif-
ferent nitrogen forms128. The characterization of tea plant
miRNA regulatory networks in response to Colleto-
trichum gloeosporioides, which is considered one of the
dominant endophytic taxa in tea plants, indicated that
miRNAs may be involved in the response to C. gloeos-
porioides attack134. miRNA characterization in tea plants
under cold and drought stresses suggested the potential
existence of an miRNA-mediated regulatory mechanism
under abiotic stresses showing coherent or incoherent
relationships with target genes to prevent tea plants from
being injured129–132. Csn-miR398a-3p-1 of tea plants was
experimentally proven to directly cleave CsCSD4, a
superoxide dismutase (SOD) gene related to the removal
of reactive oxygen species (ROS), and the expression
pattern of csn-miR398-3p-1 is opposite that of CsCSD4
under cold treatment135. Roles of miRNAs have also been
found in the bud dormancy of tea plants. Cs-miR139c
regulates the release of tea plants from bud dormancy by
suppressing the expression of its target gene, CsnTCP2
(Teosinte branched/Cycloidea/Proliferating cell factor
2)136. There is no doubt that the investigation of miRNAs
provides a broad understanding of the posttranscriptional
regulatory networks between protein-coding and
ncRNAs; however, most current progress is limited to the
identification, but not the functional verification, of
miRNAs, which will require further investigation in tea
plants.
In addition to miRNAs, long noncoding RNAs
(lncRNAs) have been also identified in tea plants using
170 RNA-seq data (~7157 million reads) from 11 tissues,
generating more than 33,000 putative novel lncRNAs137.
The tea plant lncRNAs showed tissue-specific expression
patterns and were suggested to be involved in major
aroma formation pathways of black tea137. circRNAs
(circular RNAs) were recently discovered as a new class of
ncRNAs that are covalently closed, single-stranded and
generated via back-splicing events138,139. Since their first
Page 8 of 19
documentation in A. thaliana138, circRNAs have been
detected in several other plants138,140–142. A total of 3174
circRNAs have been characterized in tea plants141. As
similarly observed in most other plants139, tea plant cir-
cRNAs exhibit tissue-specific expression patterns, and
their expression shows a positive relationship with that of
their parental genes141. Tea plant circRNAs were found to
exhibit potential miRNA-binding sites and to display
crucial functions in the photosynthetic machinery and
metabolite biosynthesis during leaf development141. In
other plants, circRNAs can act as miRNA sponges under
tomato chilling injury142, dehydration stress in wheat143,
and infection with the bacterial canker pathogen in
kiwifruit144. These studies have increasingly complicated
our understanding of ncRNAs and their regulatory
mechanisms in tea plants. Additionally, none of these
identified ncRNAs (miRNAs, lncRNAs, and circRNAs)
have been functionally well-verified regarding their roles
in regulating posttranscriptional processes in tea plants.
Further efforts aimed at accurate lncRNA characterization
and the functional examination of their interaction
mechanisms in competing endogenous RNA (ceRNA)
networks should receive more attention to elucidate their
roles in response to stress and the regulation of tea plant
secondary metabolite formation.
Genetic diversity and population structure of tea
plants
In the thousands of years since the tea plant was first
discovered, recorded, and cultivated to produce tea in
China, it has spread to more than 50 countries around the
world in tropical and subtropical regions1 (Fig. 2a). The
wide range of growing areas, long history of cultivation,
self-incompatibility and allogamy of tea plants make these
cultivars highly heterogeneous, resulting in high diversity
in both their genetics and morphology145,146. As the key
component of crop genetic improvement, the available
germplasms and the genetic diversity within a gene pool
determine the success of breeding programs. The devel-
opment of molecular markers, such as RAPD, AFLP, SSR,
and SNP markers, has allowed the successful utilization of
these markers to estimate the genetic diversity and
determine the phylogenetic relationships of different tea
germplasms147–153. Early in 1995, the systematic assess-
ment of the genetic variability of three major types (CSA,
CSS, and Cambod type) of tea plants from Kenya and the
UK suggested that the Cambod population possessed the
lowest diversity153. The further investigation of genetic
and morphological traits revealed consistent findings in
populations from other major tea plantation coun-
tries145,147,154–158. Germplasm from the Korean popula-
tion shows higher diversity than that from Japan due to
more extensive plantations in Korea and longer, intensive
tea selection programs in Japan157. The genetic diversity
Xia et al. Horticulture Research (2020)7:7 Page 9 of 19

NORTH
AMERICA ASIA
EUROPE

Tea production SOUTH AFRICA

AMERICA

0 (tonnes) 2500000

B
TPIA QTL mapping

Platform
TMDB BSA

TBC2Health WGCNA
Technology

TBC2target Transgenic technology

Germplasm Resource
Fig. 2 Global tea production and platforms/techniques for scientific studies of tea. a Global tea production across the majority of tea-
producing countries. The data were collected from 2017 FAO statistics (http://www.fao.org/). b Resource-centered research platforms and
technologies for tea molecular biology.

of the Chinese, Indian, and Sri Lankan populations is
higher than those from other countries151. Most Italian
tea plants exhibit a close relationship with cultivars from
Zhejiang Province159. Kenyan tea plants show the highest
diversity among all African germplasms, while the lowest
is found in South Africa160,161. CSA was indicated to have
donated the largest amount of genetic material in African
tea breeding161. Narrow genetic diversity is detected in the
Sri Lankan population, which is dominated by Assam-
type tea162. The estimation of worldwide tea plant genetic
diversity has increased our understanding of the popula-
tion structures and origins of tea plants, which will benefit
the global tea industry.
China hosts abundant tea plant germplasms. A wide
range of genetic diversity and population studies have
been undertaken, particularly for Chinese germ-
plasms148,149,152,163–167. Most of the Chinese tea germ-
plasms can be clustered based on their geographical origin
and genetic background149. Higher levels of genetic
diversity are observed in Guangxi, Yunnan, and Guizhou
Provinces148,167. Compared to the wild tea plants, such as
C. taliensis, the cultivated tea plants display higher
heterozygosity152.
Molecular markers are useful and are commonly
employed in crop breeding. Traditional experimental
screening of marker polymorphism is laborious and time-
consuming, especially for the broadly used SSR markers.
The development of the CandiSSR pipeline has facilitated
the efficient identification of polymorphic SSRs
(PolySSRs) based on high-throughput sequencing data,
and this pipeline has successfully identified 450 PolySSRs
in the transcriptomes of four Camellia species168. Despite
the diverse molecular and morphological markers devel-
oped, more efforts are still needed to develop extra-robust
markers (e.g., SNPs) to better facilitate the conservation
and utilization of global tea resources.
Xia et al. Horticulture Research (2020)7:7
Platforms and tools for tea plant functional
genomics
Databases
Biological databases provide scientists with an opportu-
nity to centrally access a variety of biological data. Various
tea plant databases have been established to help the tea-
related research community to better understand the
metabolome, health benefits, and genomics of tea169–171
(Fig. 2b). Yue and colleagues constructed the first com-
prehensive manually curated Tea Metabolome Database
(TMDB)171, which contains over 1400 metabolites and 600
NMR datasets collected from 364 publications. Retrieval
from the TMDB can thoroughly access a total of 24 basic
attributes of tea compounds of interest, such as compound
structure, molecular weight, and compound uses. The
beneficial effects of tea are principally derived from diverse
bioactive compounds. The establishment of the searchable
TBC2health database has connected the relationships
between tea compounds and their health beneficial effects
well169. The current release of TBC2health compromises
497 bioactive tea compounds and their associated health
effects on 206 diseases. The user-friendly response interface
contains multiple easily visualized results, aiding in the
efficient discovery of potential mechanisms of the health
benefits of tea. Increasingly, studies have confirmed that the
health benefits of tea are primarily achieved through the
regulation of target gene expression or protein activ-
ities172,173. More recently, the TBC2target database was
developed to provide candidate target genes for a total of
240 bioactive tea compounds170. The target genes were
predicted using PharmMapper via a pharmacophore-
mapping strategy174. With TBC2target in hand, scientists
can explore the potential genes targeted by tea compounds
and then reveal the possible health-promoting mechanisms
of bioactive tea compounds.
Tea plants are rich in secondary metabolites that
essentially determine tea quality2. Hundreds of studies on
tea quality and physiology have been reported, which have
generated a wide variety of specific biological data1,2. The
recently developed Tea Plant Information Archive (TPIA)
database employs the published tea plant genome as a
basic framework and incorporates a wide range of publicly
available genomic, transcriptomic, and metabolomic data
of tea plants together with globally collected tea germ-
plasms175. It also comprises over 70 transcriptomes widely
collected from 21 Camellia species, plentiful gene
expression data from various conditions (across species,
stresses, and hormone treatments), orthologs, transcrip-
tion factors, SSRs, metabolites (mainly catechins, thea-
nine, and caffeine), correlations, and manually curated
functional genes. Through long-term maintenance and
timely updating with novel datasets, the TPIA is gradually
becoming the central gateway for tea communities to
investigate the biology of tea plants in more detail, thus
Page 10 of 19
benefitting the sustainable development of the global tea
industry.
Quantitative trait loci (QTL) mapping and bulked-
segregant analysis (BSA)
QTL mapping is an efficient tool for revealing the
molecular basis of complex traits of organisms176. Despite
the fact that QTL mapping is widely used in several
crops177,178, the application of this method to tea plants is
still challenged, mainly due to the self-incompatible nat-
ure of tea plants, which exhibit a low seed yield, leading to
the propagation of insufficient populations for QTL
mapping. Nevertheless, considering the importance of tea
plants in global tea consumption, increasing efforts are
being focused on the QTL mapping of several agronomic
traits of tea plants, particularly tea yield20 and secondary
metabolites22,27–29 (Fig. 2b). A total of 23 putative QTLs
associated with tea yield were detected using 5-year yield
data from 42 F1 individuals collected across two sites in
Kenya20. These efforts led to the first QTL-mapping
investigation in tea plants, but there was no shared QTLs
discovered at the two sites, indicating an overestimated
QTL effect due to insufficient mapping populations. In
contrast to yield traits, QTLs related to tea quality have
been widely explored. Using 2-year catechins data from
183 individuals crossbred from two tea plant varieties
with distinct catechins composition, a total of 25
catechin-related QTLs were identified27. Nine of these
QTLs were validated across years and found to cluster
together in the chromosomal regions of LG03 and LG11,
suggesting a potential tandem duplication origin of tea
plant catechin genes. Similarly, 10 QTLs were found to be
involved in the determination of theobromine and caf-
feine contents using 148 individuals from a pseudo-
testcross population22. With the exception of one QTL
related to caffeine content, which was validated across
years and mapped on LG01, the other QTLs were vali-
dated from only 1 or 2 years of data, possibly due to the
relatively small size of the mapping populations. With the
expansion of mapping populations, a recent QTL map-
ping study detected several novel QTLs related to flavo-
noids29. Among the 27 identified QTLs associated with
flavonoids, 7 were newly revealed to be involved in the
determination of anthocyanin content and young shoot
color. Interestingly, quite different from other recent
reports22, QTLs controlling caffeine content were identi-
fied on LG11 in this study29. This implies that the caffeine
content of tea plants is probably controlled by multiple
genes located on different chromosomes or that the QTL
effects have been overestimated because of the limitations
of the mapping populations. In addition to tea quality and
yield traits, QTL mapping has been applied to agronomic
traits such as spring bud flush and leaf size (e.g., mature
leaf length, width, and shape indexes)28. In the future,
Xia et al. Horticulture Research (2020)7:7
efforts will be needed to further increase the mapping
populations and improve the density of genetic linkage
maps to precisely identify QTLs in tea plants.
Although over 80 QTLs have been detected for diverse
agronomic traits of tea plants, it is still difficult to explore
the candidate genes and/or alleles related to QTLs further
owing to the insufficient genetic markers and genomic
information available. The recently developed BSA
method selects and pools samples from individuals with
extreme phenotypes from biparental segregation popula-
tions and enables the efficient identification of genes and
alleles governing complex traits through statistical geno-
mic and phenotypic analyses. In tea plants, the utilization
of the BSA method coupled with RNA sequencing has
facilitated the discovery of a flavonoid 3′,5′-hydroxylase
(F3′5′H) gene that is significantly associated with catechin
content based on individuals from the two tails of an F1
population that segregates with catechin content179.
Weighted correlation network analysis (WGCNA)
WGCNA is of particular importance for tea plants to
establish the relationships between gene expression and
secondary metabolites that determine tea quality180,181
(Fig. 2b). It is also helpful for identifying clusters of highly
correlated genes and revealing their regulatory networks
in tea plant development182,183. The investigation of the
expression profiles of tea plants at 10 developmental
stages identified a total of 35 coexpression modules using
the WGCNA method with differentially expressed genes
(DEGs)182. Among these, 20 modules were found to be
related to the biosynthesis of catechins, theanine, and
caffeine, which were biologically coregulated by several
hub genes, suggesting a potential network of coordinated
regulation in three characteristic secondary metabolic
pathways of tea plants. Building the association between
tea aroma accumulation and gene expression using the
WGCNA method remarkably revealed 13 transcription
factor genes that are probably involved in terpene meta-
bolism181. Among these genes, CsOCS2 was validated
in vitro to function in terpenoid biosynthesis, indicating a
robustness and reliability of WGCNA for trait-associated
gene discovery. Similarly, the application of WGCNA
identified a gene module consisting of eight genes sig-
nificantly associated with the biosynthesis of cyanidin 3-
O-glucoside and petunidin 3-O-glucoside, helping to
elucidate the mechanism of pigmentation in the
anthocyanin-rich tea plants180. These findings suggested
that, compared to several previous correlation stu-
dies1,2,60, WGCNA is much more powerful; however, the
sample sizes and/or traits used in most of the current
studies are still limited, which decreases the accuracy and
scope of the acquired candidates. The integration of more
samples and traits from diverse cultivars and conditions in
the future will improve the capability of WGCNA and
Page 11 of 19
thus enhance the rapid discovery of candidate genes
associated with important agronomic traits in tea plants.
Future challenges and perspectives
Completion of the tea plant genome with advanced
technologies
Tea originated from China and has spread to over 160
countries around the world. According to statistics from
2017 from the Food and Agriculture Organization of the
United Nations (FAO; http://www.fao.org/), the global
tea plantation area has exceeded 4.08 million hectares,
and global total tea production has reached 6.10 million
tons, with an average increase of 0.33 million hectares
and 0.58 million tons over the past 5 years (Fig. 2a). In
stark contrast to the flourishing development of the
global tea industry, research on the major fundamental
biological problems encountered in tea plants is still
lagging behind, resulting in a low breeding efficiency
and few excellent varieties. The rapid development and
application of genomics approaches have effectively
promoted the breeding programs of crops; however,
genomic investigations of tea plants are still limited and
challenging. Although the genome sequences of two
major varieties (CSS and CSA) of tea plants were gen-
erated recently, their quality and completeness still need
to be improved1,2. Given the limitations of current NGS-
based approaches related to highly repetitive and het-
erozygous genomes, advanced single-molecule-
sequencing technologies (e.g., PacBio SMRT and
Oxford Nanopore) will help resolve the highly repetitive
regions of the tea plant genome. With the further
inclusion of linkage map and Hi-C technology to
interactively address heterozygous genomic regions, the
eventually produced chromosome-level reference gen-
ome of tea plants will facilitate comparative genomics,
evolutionary and population genetics studies and, thus,
promote breeding programs in the future (Fig. 3).
Towards tea plant pan-genomics and the phylogeny of
genus Camellia
Tea plants are widely distributed in tropical and sub-
tropical regions around the world184. They contain rich
and unique characteristic secondary metabolites, such as
catechins, theanine, and caffeine, which are essential to
the formation of tea quality. However, the contents of
these secondary metabolites vary greatly among different
varieties and Camellia species, leading to large differences
in tea processing suitability1,185. They also differ sig-
nificantly in several morphological traits (e.g., leaf size)
and stress resistance characteristics (e.g., cold tolerance),
showing a divergent genetic makeup50,186. Therefore, the
genome sequence of a single individual of a tea plant
variety (e.g., shuchazao or yunkang #10) cannot represent
the entire gene pool of tea plants. Pan-genome sequencing
Xia et al. Horticulture Research (2020)7:7 Page 12 of 19

CsC4H; CsSAMT1; CsF3H; CsGH1BG1 Genome editing

Morphology and molecular and transgenic system
phylogeny construction CsICE1 CsANR1/2 TBC2Health Improved genome
CsCBF1 CsGT1/2 CsLAR and gene annotation
Germplasm collection
CsF3'H CsLIS/NES; CsTSI
and diversity evaluation Molecular design
CsCOR1 CsTLP GWAS breeding
Cytogenetics TMDB TBC2Health Methylome TPIA

Phase I. Tea plant morphology: Phase II. Tea plant transcriptome and genome: Phase III. Tea plant population:
classification, genetic diversity sequencing, gene discovery, database construction genotype-to-phenotype mapping

2000 2005 2009 2011 2013 2015 2017 2018 2019 2030 Year

C. azalea
Revision of CSA CSS Genome sequence
CSS C. taliensis
Camellia Csβ-PD of close species
species Initiation of genome C. reticulata
C. oleifera C. japonica MYB75 Knowledgebase for
CsTCS sequencing projects
C. chekiangoleosa C. petelotii CsGSTF1 tea researchers
C. sasanqua CsUGT85A53
Transcriptome Genome Database Key functional gene

Fig. 3 Timeline of research on tea plant genetics and genomics. The solid black circles indicate past events in tea plant genomics, including
Phase I of tea plant morphology and Phase II of tea plant transcriptome and genome studies. Events are highlighted with colored rectangular boxes:
yellow (transcriptome sequencing), orange (genome sequencing), cyan (database development), and gray (gene cloning). The solid white circles
represent Phase III of tea plant comparative genomics and population genetics studies in the future.

is a newly developed strategy for investigating the genetic
variations among different subspecies and/or individuals
that has been widely used in several crops and model
species, such as rice187, soybean188, maize189, and Arabi-
dopsis190. With the completion of the tea plant genome,
more efforts are still needed to further investigate the
pan-genomics of tea plants from different varieties and/or
ecotypes to expand the tea gene pool. However, the
phylogeny of genus Camellia and the relationships among
different tea populations remain largely unclear, which
has seriously hindered the in-depth exploration of pan-
genomics in tea plants.
Although several studies have attempted to resolve the
phylogeny of genus Camellia using chloroplast and/or
mitochondrial DNA sequences, the generated phyloge-
netic trees are somewhat poorly supported, primarily
due to the high frequency of hybridization and poly-
ploidization, as well as the relative conservation of
organelle DNA sequences among Camellia spe-
cies39,45,191–193. Compared to organelle fragments,
massive nuclear gene sequences rapidly obtained from
genome and transcriptome sequencing provide abun-
dant candidate low-copy-number nuclear genes and
informative sites for phylogenetic investigation and have
been increasingly applied to clarify the phylogenies of
some complex plant taxa194–198. Therefore, the
sequencing of additional genomes and/or tran-
scriptomes of Camellia species, as well as elite tea cul-
tivars in the foreseeable future will not only help to
reveal their complex phylogenetic relationships but will
in turn promote pan-genomic studies of tea plants to
expand the gene pool.
Genome-wide association study (GWAS) of tea plants
GWAS is a widely accepted strategy to identify genes/
alleles associated with complex traits in crops with the
rapid innovation of NGS technologies199–202. Genotypes
(typically based on SNPs) and phenotypes (usually those
of different traits) are the two fundamental requirements
in GWAS, in which a higher SNP density and more
diverse phenotypes of a population will theoretically
produce more reliable and significant associations. How-
ever, conventional GWAS design and methods have
resulted in challenges in the identification of multiple
functional alleles within a single gene as well as rare alleles
in the population203,204. GWAS of tea plants has also been
confronted with major difficulties in population con-
struction and phenotyping related to germplasm collec-
tion and the ultraslow growth of tea plants when
transplanted to nursery gardens. Nevertheless, consider-
ing the high level of genetic and morphological diversity
in tea plants, GWAS is still the most efficient way to
detect candidate loci of complex traits undergoing
improvement. To apply GWAS to study the genetics
underlying tea traits of interest, the following goals should
be considered: (1) collection and construction of core
germplasms with adequate genetic diversity; (2) trans-
plantation of the germplasms to one or more local gar-
dens for accurate multiple-year phenotyping; (3) further
improvement of the quality and annotation accuracy of
the current genome assembly; (4) integration of GWAS
data with other omics datasets (e.g., transcriptomic and
metabolomics data); and (5) construction of an efficient
transgenic system to functionally validate the candidates.
In this way, GWAS can be effectively applied to
Xia et al. Horticulture Research (2020)7:7
understand the genetic basis of agronomic traits asso-
ciated with tea quality, thus enabling the molecular design
of new cultivars in the near future.
Origin and domestication of tea plants
China is documented as the first country to cultivate
and utilize tea plants in the world205. It harbors abundant
tea germplasms and has long been considered the origin
of tea plants206. However, since no wild ancestor of tea
plants has been discovered in China, the origin and
domestication of tea plants are still a mystery and attract
worldwide attention. Although recent progress has been
made in understanding the origin and domestication of
tea plants based on molecular markers (primarily SSRs),
the resultant conclusions are unilateral and even con-
troversial due to the poor representativeness of sample
collections and the lack of sufficient molecular markers
for diversity evaluation160,161,207–211. The recent release of
tea plant genome sequences has laid a solid foundation for
solving this problem1,2; however, further efforts focusing
on the following issues are still needed: (1) collection of
globally representative tea plant samples; (2) investigation
of the population structure of tea plants and identification
of their putative wild ancestor; (3) estimation of popula-
tion diversity using genome-wide SNP markers; (4)
scanning of genomic regions with significantly lower
diversity in cultivated but not wild tea plants and identi-
fication of the candidate genomic regions selected during
domestication; and (5) functional investigation of the
characteristics of genes within regions of domestication. It
should be noted that most of the existing tea varieties are
bred and propagated directly from natural populations.
Artificial domestication may have had little impact on the
variation in genome sequences. Therefore, tea plants may
actually undergo adaptive evolution compared to other
crops (e.g., rice) subjected to strong domestication
pressure212.
Establishment of high-efficient transgenic system
Agrobacterium-mediated genetic transformation has been
extensively applied to verify the function of genes with the
aim of improving the quality and yield of crops. In tea
plants, the first transgenic plant was generated using
Agrobacterium-mediated transformation in somatic
embryos213. However, at least 45 weeks were possibly
needed for the transformation procedure alone, and addi-
tional years were required for further transplantation213. It
also required several years of experimental work to establish
stability and germline transmission in these plants.
Although several efforts have been made to optimize the
transformation system of tea plants using different strate-
gies214–223, transformation efficiency still represents a
challenge under the available methods215,224. In the face of
these challenges, some key points related to transgenic
Page 13 of 19
technology need to be further detailed: (1) optimization of
the transformation and regeneration system; (2) clarifica-
tion of the transformation mechanism of tea plants, parti-
cularly to elucidate the effects of tea polyphenols on
Agrobacterium infection; and (3) identification of more
suitable tea varieties for better transformation and regen-
eration. The most recent CRISPR system has enabled effi-
cient genome editing in crops and shows great potential for
accelerating precise crop improvement225. Considering the
difficulties encountered transgenic studies of tea plant, the
establishment of the CRISPR system will promote the
transgenic progress in tea plants221. More recently, scien-
tists demonstrated an efficient approach to overcome self-
incompatibility in diploid potato using the CRISPR/
Cas9 system through S-RNase locus knockout, which pro-
vided a useful example for the further application of
transgenic technology and breeding in tea plants, con-
sidering their similar self-incompatibility features226.
Collection and conservation of tea plant germplasms
Resources are clearly the key to crop genetic improve-
ment. The success of plant breeding and conservation is
largely dependent on the amount and distribution of
genetic variations present in collections. Currently, tea
germplasms are becoming the most valuable fundamental
material for tea breeding and biotechnology studies, pre-
senting great potential in the whole tea industry227. The
remarkable achievements made in worldwide tea plant
germplasm surveys, collection, and conservation have
preserved more than 15,000 tea plant accessions accord-
ing to incomplete statistics from the China Tea Science
Society (http://www.chinatss.cn/). These invaluable
germplasms have driven the extraordinarily rapid devel-
opment of tea plant genomics, genetics, and breed-
ing151,152,175,207,228,229. However, most of the current
accessions that have been collected and used are varieties,
and few “wild” or close relatives have been integrated.
Another major problem that is currently faced by the field
is the imbalanced utilization and protection of cultivars
versus “wild” germplasms. The survival environment of
“wild” tea resources has been constantly destroyed with
rapid economic development and urbanization. In addi-
tion, local germplasms (landraces) with specific char-
acteristics are on the edge of being lost due to the
popularization of elite varieties in the past few years.
Above all, the surveying, collection, and conservation of
tea plant germplasms should be urgently emphasized.
Otherwise, no diverse tea plant resources or only a few
mono varieties will survive and be available for utilization
for the genetic improvement of tea in the future. We
therefore propose that a tea plant conservation alliance
uniting the global tea-planting countries needs to be
established to accelerate the preservation and utilization
of global tea plant resources.
Xia et al. Horticulture Research (2020)7:7
In-depth understanding of the complex secondary
metabolism network
The overall importance of tea plants as an economic
crop is due to the health-promoting functions of teas as
non-alcoholic drinks that are popular worldwide230.
Numerous studies have documented the health benefits
of various tea drinks in humans, and studies continue to
produce more fundamental discoveries about the effects
of functional tea components on the improvement of
human heath231,232. It is not only the three major types
of tea plant secondary metabolites (catechins, caffeine,
and theanine) but also the volatile terpenoids, saponins,
polysaccharides, and other phenolic conjugates found in
tea that contribute to the beneficial health effects and
the enjoyable flavors of various teas70,71,233. As men-
tioned above, our understanding of these complicated
secondary metabolites in tea plants in terms of their
biosynthesis, transport, and storage as well as their
regulation is very limited. This is one of the major
obstacles to the breeding of high-quality tea plant
varieties with biofortified nutrients or functional com-
ponents or other special properties. Future work could
focus on basic questions about tea secondary metabo-
lism, which will facilitate the breeding of tea plants with
desirable qualities and benefit human health improve-
ment projects.
In conclusion, tea plants are perennial woody plants in
nature with a long growth cycle, which severely limits
their genetic breeding. Traditional cross-breeding is
extremely difficult and time-consuming for tea plants
because most of the existing tea varieties are bred and
propagated directly from natural populations. Modern
transgenic breeding technology has provided us a new
solution for the molecular design of breeding strategies,
which basically relies on the development of functional
genomics and molecular biology. Although great progress
has been made in the last two decades, the genomics and
molecular biology of tea plants are still not fully under-
stood, mainly due to difficulties in resource collection and
identification, the generation and identification of mutant
plants, and population construction. In particular, the
limited knowledge of the functional genomics and
developmental biology of tea plants has narrowed our
understanding of their basic biological characteristics.
Therefore, compared to other crops such as rice, there is
still a long way to go in tea plant genomics and molecular
biology research. In the near future, scientists should
focus more on industry-oriented major basic biological
research based on germplasm collection and utilization,
particularly by deciphering the tea plant genome, as an
opportunity for enhancing studies on the mechanisms of
the biosynthesis and regulation of secondary metabolites,
the genetic basis of important agronomic traits, the
molecular mechanisms of stress and disease resistance,
Page 14 of 19
and the developmental biology of tea plants, to accelerate
the molecular design breeding program and, thus, pro-
mote industrial development.
Acknowledgements
We would like to thank Dr. Penghui Li, Dr. Zhaoliang Zhang, and Dr. Chuankui
Song for his discussion and comments on the manuscript. We apologize for
not being able to cite many of the excellent publications on tea plants due to
the limit of paper length. The authors acknowledge support from the National
Natural Science Foundation of China (31800180), the Natural Science
Foundation of Anhui Province (1908085MC75), the National Key Research and
Development Program of China (2018YFD1000601), the Science and
Technology Project of Anhui Province (13Z03012), the Special Innovative
Province Construction in Anhui Province (15czs08032), the Changjiang
Scholars and Innovative Research Team in University (IRT1101), the China
Postdoctoral Science Foundation (No. 2017M621992), and the Postdoctoral
Science Foundation of Anhui Province, China (No. 2017B189).
Conflict of interest
The authors declare that they have no conflict of interest.
Supplementary Information accompanies this paper at (https://doi.org/
10.1038/s41438-019-0225-4).
Received: 10 July 2019 Revised: 17 October 2019 Accepted: 3 November
2019
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